34] Allotopic expression of yeast mitochondrial maturase to study mitochondrial import of hydrophobic proteins

Abstract

Allotopic expression of a gene implies that a new copy of a gene, localized to a new cellular compartment, for instance, from mitochondria to the nucleus, is able to confer the corresponding wild-type phenotype. Thus, in the case of the allotopic expression of a mitochondrial gene, the corresponding cytoplasmically translated product should be targeted, in an active form, to the adequate mitochondrial subcompartment. This chapter describes the methods to transfer a mitochondrial gene to the nucleus. It illustrates the example of the bI4 RNA maturase to show how a cytoplasmically translated form of this protein can complement RNA maturase mitochondrial mutations. The chapter also explains how the RNA maturase can be used as a sensitive reporter activity, to study the mitochondrial import of hydrophobic proteins, and how this can permit the development of a genetic screen to search for genes involved in this import process.